Antiviral effect of siRNA D3 in relation to the infectious dose of Langat.

<p>HeLa cells were infected with Langat virus at different infectious dose (MOI of 10, 1, 0.1 or 0.01) and one hour after infection, cells were treated with 200 nM siRNA sequence D3 or with nonsense siRNA. Virus titer was assessed by quantitative real-time RT-PCR 6 days after infection in the cell culture supernatant. The reduction of viral RNA in siRNA D3 treated cells is statistically significant compared to nonsense siRNA treated cells for all four MOI (p<0.05). The data are presented as mean ± SD of three independent experiments and significance calculated using an unpaired T-test.</p

Inhibitory effects of siRNA D3 on viral replication on HeLa cells over time.

<p>The replication profile for Langat virus was determined for the time period of 6 days. Indicated are the numbers of RNA copies (A) and the number of infectious particles (B) for cell cultures treated with nonsense siRNA or specific siRNA D3.</p

LGTV-specific siRNA sequences inhibit production of viral RNA in HeLa cells.

<p>Nineteen siRNA sequences (Q1–Q6 and D1–D13) targeting genes within the whole open reading frame of LGTV genome were analyzed for their antiviral potential on HeLa cells (A). After transfection with siRNA, cells were infected with Langat virus (MOI = 10) and six days later virus replication was assessed by quantitative real-time RT-PCR (B). Results are shown as a percentage of virus inhibition compared to the control cells transfected with the non-coding siRNA. Data are presented as mean ± SD of three independent experiments. Fourteen (Q2, Q3, Q4, Q5, Q6, D3, D4, D5, D6, D8, D9 D11 D12 and D13) out of 19 were significantly reduced (p<0.05) compared to a theoretical mean of 100% expressed by cells treated with nonsense siRNA; measured by One sample t test.</p

Learning capacity was assessed in the Morris water maze after 4 training days.

<p>In probe trials, the mean distance of the animals to the previous location of the platform was calculated. LiCl led to a significantly improved learning capacity compared to NaCl on day 5 (2way ANOVA). In post-hoc analysis, the effect of LiCl treatment was significant in infected animals (PM+) while it remained below statistical significance in mock-infected animals (PM−). (Boxes extend from the 25th to 75th percentiles and include median; +, mean; whiskers, minimum to maximum value; *p<0.05).</p

Cortical damage was quantified 42 h after induction of bacterial meningitis in cryosections.

<p>(<b>A</b>) LiCl treatment reduced cortical injury without reaching statistical significance when compared to littermates treated with NaCl. (<b>B</b>) The effect was below statistical significance when comparing animals with lithium serum concentrations ≥0.4 mmol/l to NaCl treated littermates. (Boxes extend from the 25th to 75th percentiles and include median; +, mean; whiskers, minimum to maximum value).</p

Cyto-/chemokine concentrations in cerebrospinal fluid samples of rats with pneumococcal meningitis and treated with NaCl or LiCl were measured 18 h after infection.

<p>All cyto-/chemokines measured were elevated in infected animals receiving LiCl compared to their littermates receiving NaCl. For TNF (<b>A</b>), IL-10 (<b>B</b>), and MCP-1 (<b>C</b>) this difference reached statistical significance. (TNF, tumor necrosis factor; IL, interleukin; MCP-1, monocyte chemoattractant protein 1; boxes extend from the 25th to 75th percentiles and include median; +, mean; whiskers, minimum to maximum value; *, p<0.05; **, p<0. 01).</p

At the time of sacrifice, lithium concentrations in serum and cerebrospinal fluid (CSF) of infected animals show a significant correlation (r = 0.91; p<0.0001; n = 15).

<p>At the time of sacrifice, lithium concentrations in serum and cerebrospinal fluid (CSF) of infected animals show a significant correlation (r = 0.91; p<0.0001; n = 15).</p

Expression of genes relevant in apoptotic pathways was analyzed 42 h after infection (PM+) or mock-infection (PM−) by qPCR.

a<p>values are mean ± standard deviation (arbitrary units), numbers of animals per group are indicated in parenthesis;</p>b<p>2way ANOVA;</p>c<p>Sidak’s multiple comparisons test;</p>d<p>n = 3; *, p<0.05; **p<0.01; qPCR, quantitative real-time polymerase chain reaction; <i>bdnf</i>, brain-derived neurotrophic factor; <i>bcl2</i>, B-cell lymphoma protein-2; <i>bax</i>, Bcl-2-associated X protein; <i>tp53</i>, tumor protein p53.</p><p>Expression of genes relevant in apoptotic pathways was analyzed 42 h after infection (PM+) or mock-infection (PM−) by qPCR.</p

Cyto-/chemokine levels were measured in cerebrospinal fluid samples of infant rats 18 h after infection.

a<p>values are mean ± standard deviation (median, min., max.);</p>b<p>lithium serum conc. 0.4 mmol/l – 1.5 mmol/l;</p>c<p>non-parametric distribution; *, p<0.05; **, p<0.01; TNF, tumor necrosis factor; IL, interleuki n; MCP-1, monocyte chemoattractant protein 1; MIP-1α, macrophage inflammatory protein 1 α; IFN-γ, interferon gamma.</p><p>Cyto-/chemokine levels were measured in cerebrospinal fluid samples of infant rats 18 h after infection.</p

Additional file 4: of Foreign peptide triggers boost in pneumococcal metabolism and growth

Table S3. RNA-Seq data for ΔORF 2 mutant with and without ORF 2 peptide. Table shows only significant changes in expression. A significant change in expression was observed for 249 genes of which 177 were upregulated by the ORF 2 peptide and 72 were downregulated by the peptide. (PDF 231 kb